Currently, the primary cause of death in Japan is cancer. In particular, it is quite difficult to prevent cancer metastasis completely with preexisting operational therapy, chemotherapy, or radiation therapy; thus, there is a need for research on newer therapies; and in particular, gene therapy is attracting attention and intensively studied. Based on basic studies, clinical tests on cancer gene therapy have been practiced frequently over past decades or so mainly in the United States as well as in Japan and other advanced countries, and the number of such patients is increasing year by year. However, there is no report that a patient was cured completely by the gene therapy. The biggest cause thereof is that most of the gene-introducing vectors currently used are nonproliferating viral vectors designed only to introduce a therapeutic gene after viral infection and not to proliferate, from the genetic engineering point of view, for safety. The nonproliferating viral vectors are indeed safer, but such a viral vector does not, of course, introduce the gene into the regions other than those into which the liquid containing the vector penetrates by in-vivo administration in actual clinical settings, even if the viral vector shows an excellent transferring efficiency in an in-vitro test using cultured cell. When a nonproliferating vector is used, it is not possible to overcome the problem of the recurrence of cancer from cancer cells in the organs where no gene is introduced, and that is the biggest cause for the gene therapy not giving expected clinical results.
There is a report of a variant adenovirus (ADV) that lacks a region in E1B that proliferates specifically in a p53 function-deficient cancer cell, which is often in the incompetent state, as an adenoviral vector (hereinafter, referred to as ADV) overcoming such a problem (Bischoff J. R., et all., Science. 1996 Oct. 18; 274 (5286): 373-376). Since then, such proliferation-regulated viral vectors, which proliferate specifically in cancer cell and do not proliferated in normal cell, have been studied. For example, an attempt was made to proliferate an adenovirus specifically in prostatic cancer cells by expressing the adenoviral E1A gene with a prostatic cancer-specific PSA promoter (Rodruguez, R., et al., Cancer Res, 57, 2559-2563, 1997).
However, proliferation-regulated viral vectors hitherto reported were designed to be regulated by a single factor, or the vectors were designed to proliferate only in cancer cells in response to a single factor by using the difference in expression of the factor in cancer cells and normal cells, to make the vectors specific to cancer. However, the difference between cancer and normal cells is not specified definitely only by a single factor, and thus, conventional proliferation-regulated viral vectors were far from specifically targeted to cancer. It would be necessary to use multiple factors different in properties simultaneously, to make the vector more specifically targeted to cancer, but there is no such a report so far. In addition, each of the traditional proliferation-regulated viral vectors should be prepared one by one separately in gene recombination. Adenovirus is a long DNA virus of 36 kB in length and thus, there are many restrictions in gene recombination thereof; for example, the number of the restriction enzymes for use in recombination of a plasmid vector containing the same is limited; and thus, it was impossible to perform the recombination efficiently. Thus, it was technically impossible to produce great amounts of proliferation-regulated adenoviral vectors rapidly and to prepare and evaluate a lot of proliferation-regulated viral vectors rapidly for use in screening vectors specifically targeted to cancer.